ABSTRACT Down Syndrome (DS), or Trisomy 21, is the leading genetic cause of developmental and cognitive disability in children. As individuals with DS live longer, we now realize most develop early-onset Alzheimer Disease (AD). In addition, people with DS have greatly increased risk of congenital heart disease, myeloproliferative disorder and leukemia, as well as immune and other system defects. Hence, understanding how three copies of normal genes on chromosome 21 impacts cell expression profiles and cell phenotypes is important not only for DS, but for conditions that afflict the non-DS population, particularly AD, perhaps the biggest medical challenge of our time. Researchers have sought to identify the transcriptional and phenotypic changes responsible for the various aspects of DS; however, this work is hampered by individual variation as well as the genetic complexity and phenotypic variability of DS. Thus, there is a critical need for better ways to determine specific cellular pathologies and gene pathways which underlie aspects of the syndrome, and to facilitate screening of drugs to correct them. We have recently created and demonstrated such a system and, supported by compelling preliminary results, propose to use it to ?dissect? the cellular impact of Trisomy 21, on genome-wide pathways and phenotypes, in undifferentiated pluripotent and in neural DS stem cells. We will capitalize upon this unique system to investigate the most direct effects of Trisomy 21 on genome-wide changes in expression of specific genes and pathways (Aim1), correlate this with specific neural cell phenotypic changes (Aim2), and ultimately examine the contribution of specific Chr21 genes (Aim 3) to DS and AD pathology. Rather than focus on one or more ?favored? aspects or hypotheses, we propose an unbiased and broad approach, supported by several collaborators who are leaders in their areas of molecular cell biology and neurobiology. This novel approach has great promise to surmount challenges that have confounded clear understanding of the basic biology of DS, and thus provide the foundation for longer-term translational efforts to treat DS. Greater understanding of genes and cell pathways perturbed in human DS is important for development of drug therapies (for DS and AD), but also has implications for the potential development of ?chromosome silencing? or gene therapies. This work also has broad basic impact for understanding genome balance, the coordinated levels of expression for genes throughout the genome.